Recirculating the cultivation medium of Spirulina platensis (S. platensis) enables efficient water and nutrient recycling, thereby reducing production costs. To figure out the inhibition components of the reused medium and cell oxidate response, this study delves into the metabolic regulation of the reused medium and its extracted organic matters (OMs) and extracellular polysaccharides (EPS) on S. platensis. The reused medium and the medium containing dissolved OMs and EPS significantly increased oxidative stress in S. platensis, reducing biomass production with inhibition rates ranging from 18.08 % to 26.59 %. Nevertheless, the incorporation of EPS from OMs augmented the synthesis of proteins, polyphenols, and chlorophyll in S. platensis, sustaining photosynthetic activity and a higher proportion of live cells. Future research should prioritize the characterization of OMs and EPS, mitigate the inhibitory effects of OMs extracted residue (molecular weight <
1000 Da), further optimize the recyclability of the reused medium, and enhance S. platensis's functional composition.